Adiponectin/Acrp30 is a recently identified adipose tissue-derived hormone with anti-diabetic and insulin sensitizing functions. How the adiponectin signal is transduced from its receptor and regulated in cells is currently unknown. To identify interacting molecules downstream of the adiponectin receptor, we screened a yeast two-hybrid cDNA library using the intracellular portion of the adiponectin receptor 1 (AdipoR1) as bait. This screening led to the identification of APPL-1, an Adaptor protein containing a Phosphotyrosine binding PTB) domain, a Pleckstrin homology (PH) domain, and a Leucine zipper motif, as an AdipoR1-associated protein. APPL-1 interacted with AdipoR1 in vitro and in mammalian cells. Importantly, overexpression of APPL-1 stimulated phosphorylation of AMP activated protein kinase (AMPK) and p38 mitogen activated protein kinase (MAPK), both of which have been shown to mediate adiponectin-stimulated fatty acid oxidation and glucose uptake. These novel findings suggest that the interaction between AdipoR1 and APPL-1, the only intracellular signaling molecule identified so far that binds to the adiponectin receptor, may be the missing link between adiponectin binding and its downstream events. To test this hypothesis, we will: 1) Define the molecular mechanisms that regulate the interaction between APPL-1 and AdipoR1; 2) Characterize the biochemical mechanism by which APPL-1 stimulates p38 MAPK phosphorylation and the functional role of APPL-1 in adiponectin-stimulated p38 MAPK activation and downstream function; 3) Elucidate the biochemical mechanism by which APPL-1 stimulates AMPK phosphorylation and characterize the functional role of APPL-1 in adiponectin-stimulated AMPK activation and downstream function. Results from these studies will not only shed light on our understanding of the adiponectin signaling pathway and its regulation, but will also provide valuable information on the design of new pharmacological interventions for clinically important diseases such as obesity and Type 2 Diabetes.